Schlafen4+-MDSC in Helicobacter-induced gastric metaplasia reveals role for GTPases

Abstract

IntroductionMDSCs express SCHLAFEN 4 (SLFN4) in Helicobacter-infected stomachs coincident with spasmolytic polypeptide-expressing metaplasia (SPEM), a precursor of gastric cancer. We aimed to characterize SLFN4+ cell identity and the role of Slfn4 in these cells.MethodsSingle-cell RNA sequencing was performed on immune cells sorted from PBMCs and stomachs prepared from uninfected and 6-month H. felis-infected mice. Knockdown of Slfn4 by siRNA or PDE5/6 inhibition by sildenafil were performed in vitro. Intracellular ATP/GTP levels and GTPase activity of immunoprecipitated Slfn4 complexes were measured using the GTPase-Glo assay kit. The intracellular level of ROS was quantified by the DCF-DA fluorescent staining, and apoptosis was determined by cleaved Caspase-3 and Annexin V expression. Gli1CreERT2 x Slfn4fl/fl mice were generated and infected with H. felis. Sildenafil was administered twice over 2 weeks by gavaging H. felis infected mice ~4 months after inoculation once SPEM had developed.ResultsSlfn4 was highly induced in both monocytic and granulocytic MDSCs from infected stomachs. Both Slfn4+-MDSC populations exhibited strong transcriptional signatures for type-I interferon responsive GTPases and exhibited T cell suppressor function. SLFN4-containing protein complexes immunoprecipitated from myeloid cell cultures treated with IFNa exhibited GTPase activity. Knocking down Slfn4 or PDE5/6 inhibition with sildenafil blocked IFNa induction of GTP, SLFN4 and NOS2. Moreover, IFNa induction of Slfn+-MDSC function was inhibited by inducing their reactive oxygen species (ROS) production and apoptosis through protein kinase G activation. Accordingly, in vivo disruption of Slfn4 in Gli1CreERT2 x Slfn4fl/fl mice or pharmacologic inhibition by sildenafil after Helicobacter infection also suppressed SLFN4 and NOS2, reversed T cell suppression and mitigated SPEM development.ConclusionTaken together, SLFN4 regulates the activity of the GTPase pathway in MDSCs and precludes these cells from succumbing to the massive ROS generation when they acquire MDSC function.

Schlafens: Emerging Therapeutic Targets

“…Previously, Ding et al demonstrated that SLFN4+ myeloid-derived suppressor cells (MDSCs) could migrate to the stomach following Helicobacter pylori infection potentiating the development to gastric metaplasia [58]. Similarly, SLFN12L (which exhibits sequence similarity with Slfn4) has been shown to colocalize with MiR130b, a microRNA that was associated with SLFN4+ MDSCs that suppressed T cells and promoted Helicobacter-induced metaplasia [59].…”

Hedgehog pathway and cancer: A new area (Review)

“…Then, these cells differentiate and mature under the induction of factors such as IFN-α, express GLI1-dependent schlafen 4 and secrete interleukin (IL)-1β to activate the IL-6/phosphorylated signal transducer and activator of transcription (STAT)-3 pathway. The activation of SHH during Hp infection was closely related to the expression of programmed death-ligand 1 (PD-L1) ( 128 ), as well as the emergence of myeloid-derived suppressor cells (MDSCs), intestinal metaplasia and soluble polypeptide expression metaplasia ( 129 ).…”